 All right, so good morning everyone. So welcome back to our hematology class for today. So we're actually finally going to discuss the last topic for the semifinals which we'll be talking about Porphyria, Cideroblastic Anemia, and Hemoglobinopathy. Please do remember that I will be skipping the intrinsic and extrinsic causes leading to destruction of RBC or hemolysis or what we call hemolysis. So that will be now part of the final examination. So for today, we're going to talk about Porphyria, Cideroblastic Anemia, and the different hemoglobinopathy. So let's get started with our discussion for today. So the first one is all about Cideroblastic Anemia. So Cideroblastic Anemia is actually characterized to be an interference in the production of your protoporphyrin ring. So remember that your protoporphyrin ring is a major component in your heme synthesis or your heme production. So Cideroblastic Anemia causes your microcytic and hypochromic anemia. And Cideroblastic Anemia is said to have a hallmark symptom or a hallmark mark, a hallmark of Cideroblast. So your ring Cideroblast is characterized by the deposition of your iron in your precursor cells in your bone marrow causing now your ring Cideroblast. So let us just try to differentiate. In Cideroblastic Anemia, I would try to compare it with your iron deficiency anemia. In your iron deficiency anemia, I hope you still remember, in your iron deficiency anemia, we did talk about of your free erythrocyte protoporphyrin. Your free erythrocyte protoporphyrin or your FEP is said to be increased in iron deficiency anemia. Bakit po ganun sir? Kasi nga marami kang protoporphyrin ring, pero wala kang iron to bind with them to form your heme. That is why in your iron deficiency anemia, there is a deposition of excess protoporphyrin. Those excess protoporphyrin now are detected using FEP. Sir, why are you trying to compare the iron deficiency anemia with regards to Cideroblastic Anemia? Dahil sa kato ni Cideroblastic Anemia, the problem this time is that you have enough iron reserve. You have enough iron inside your body, but the problem this time is that there are no enough protoporphyrin to bind this time on your iron. Nakukuha, this time siya Cideroblastic Anemia, ang kulang naman natin is yung ating protoporphyrin ring or protoporphyrin 9 natin. That is why walang magkukombine with your ferrous iron to form your heme. In that case, the deposition now of those excess iron in your mitochondria will now lead in the formation of your Cideroblastic cells or rather your ring Cideroblastic. So your ring Cideroblastic is the deposition of those iron on your mitochondria. And maybe some of you would be wondering, sir, what is now the difference between your Cideroblastic Anemia, your hemochromatosis, and your hemociderosis? They are different because in Cideroblastic Anemia, there is no iron overload. Wala tayong iron overload in Cideroblastic Anemia. It's just that the iron that is supposedly to be used in the production of your heme eventually your hemoglobin was not used. That's why it accumulated on your mitochondria. And did you know that Cideroblastic Anemia are usually acquired? And one of the leading cause of acquired Cideroblastic Anemia is your lead poisoning. So you all know your lead poisoning or your plumbism in your RBC anomalies. You did mention about lead poisoning causing your basophilic stippling, correct? Causing your basophilic stippling. So it causes basophilic stippling because it inhibits a enzyme that clear out your RNA remnants. But it's just more than the enzyme that is being inhibited there. Aside from that enzyme, it also inhibits key enzymes in the production of your heme. When lead poisoning or plumbism is happening, you would also expect that there will be an interference in the production of your proto-parfarin ring. And how does your lead affects your proto-parfarin? So your lead interferes in two ways. And in these two ways, it affects the two major processes in your heme synthesis. And what are those? This is the conversion of your aminolevolinic acid or your ala into becoming your porphobilinogen or your PBG. So it inhibits your ala dehydratase or your PBG synthase. So this results now into the accumulation of your aminolevolinic acid. In addition to that, your lead also interfered in the process of incorporating your ferrocylion into your proto-parfarin 9. And in this case, the enzyme being inhibited is your ferrokylates or your heme synthase, which then result in the accumulation of iron and proto-parfarin in the mitochondria. Thereby leading in the formation of your ring, cederoblast. So again, for one more time, your lead interferes your ala dehydratase or your PBG synthase. And it also interferes your ferrokylates and your heme synthase. So laging yung katandaan, like what I always say, kapag yung enzyme is inhibited, there will be an accumulation of its substrate. So in this case, it will be an accumulation of its substrate. But I always say, kapag yung enzyme is inhibited, there will be an accumulation of its substrate. So in this case, dahil wala ng ALA dehydratase or PBG synthase, tataas si ala bababa, si porfo bilin na siyan. At the same time, in this case, tataas naman din si proto-parfarin 9 at bababa yung formation ng ating heme. And dahil mababa ang formation ng yung heme, wala mapuproduce na hemoglobin, thereby causing now your cederoblastic anemia, which is a microcytic hypochromic anemia. One thing that is also very much associated with cederoblastic anemia is your porphyria. Porphyria, on the other hand, is more on the hereditary condition that impairs your heme synthesis. Or impairs your proto-parfarin production. That is why we call it porphyria. Porphyria is the accumulation and deposition of porfarin in your hepatocytes. In your hepatocytes and even in your cells. That is the reason why accumulation or leakage of your porfarin will now lead to the leak of porfarin into your cells as they age and die. And this can also be excreted in your urine and feces. By the way, this porfarin that are accumulated in your body, or in your body tissue, would also cause fluorescence in some of the body parts that are being deposited with porphyria. For example, your teeth and your bone will fluoresce because of porphyria. So luminos yung nipin mo, imagine that. Luminos yung nipin mo. At the same time, psychosis is also a very prominent clinical feature, especially during the past century, which were seen in intermarrying European monarchy. At the same time, psychosis is also a very prominent clinical feature, especially during the past century, in European monarchy in Europe before. So naki kita si psychosis. So in addition to that, because of the deposition of your porfarin now in your skin, this can also lead photosensitivity when exposed to sunlight. So your porphyria can come into three different forms which are all hereditary by nature, which are your congenital erythropoietic porphyria, or so-known as your Gunter's disease. Your erythropoietic protoporphyria and your ex-linked erythropoietic protoporphyria. So all of this would actually show photosensitivity. This is normalcytic normochromic. This would cause mild anemia and this would cause mild microcytic hypochromic anemia. So all of these three are different from one another because of the enzyme that is deficient. So in here, the one deficient is your uroporferinogen 3 synthase, this one that there is a deficiency in your ferrocheletase or your hemsynthase and here there is a deficiency in your ALA synthase. ALA synthase. If you would actually notice, if you would actually notice, porphyria is actually the cost or the reason of your folklore about your vampires. Your folklore about your vampires because they are photosensitive and some of them have psychosis. Some of them have psychosis. So also in night, dahil nagfluoflores ang tit nila, you would really see that these patients are actually the one that is dubbed as to being vampires. Siling mga tinatawad na vampires during that time. But in reality now, we all know that it's actually not vampires but actually porphyria. That is actually just your porphyria. So please do remember that. Please do remember that. So moving forward, let's go on now to your hemoglobinopathy. So your hemoglobinopathy. So hemoglobinopathy refers to the disease states involving now pure hemoglobin molecule. Kanina, we just talked about hem. Kung mapapansin ninyo, yung progression ng pagde-diskas ko sa inyo, last time we talked about iron deficiency anemia. So may problem sa iron. Ngayon naman may problem tayo sa hem kaya nakaroon tayo ng sideroblastic anemia and even your porphyria. Now the problem is the hemoglobin itself. So hemoglobinopathy refers to the disease state involving your hemoglobin molecule. So hemoglobinopathy are the most common genetic. This is affecting approximately 7% of the world's population. The world's population. So for this morning, I will only be discussing about hemoglobinopathy but it is just right for me to differentiate hemoglobinopathy in your talasemia. Kasi baka nakakaroon kaya nang baka later on, you'd actually be confused with this two term. To be specific, talasemia is actually considered to be an a-hemoglobinopathy as well. It is also a-hemoglobinopathy. But the difference between hemoglobinopathy the difference why may hiwala tayo na talasemia is because qualitative hemoglobinopathy are called your structural defect hemoglobinopathy. So this is due to altered amino acid sequence. So dahil may mutation sa DNA natin kapag ito ay na translate into becoming your protein or your becoming your amino acid or your protein there will now be alteration in the amino acid sequence. This altered amino acid sequence would cause now alteration in their structure causing now structural defect and alteration in their function. That's why we call this collectively as your qualitative hemoglobinopathy. May problema sa quality pero tama lang yung production unlike your talasemia. Your talasemia is characterized to have a reduced rate of hemoglobin synthesis. That's why it is your quantity that's why it is your quantitative hemoglobinopathy. Your quantitative hemoglobinopathy na hukuha. So talasemia normal yung structure normal yung function it's just that depleted yung production depleted yung production o hindi mababa yung production sa ganyong type ng hemoglobinopathy. Two types of hemoglobinopathy. I hope those two are clear. So qualitative ang problema sa quality ng hemoglobin we call it your structural defect or qualitative hemoglobinopathy. But if it is your quantitative now the quantity of hemoglobin is the one depleted we call it talasemia. So before we go deeper into the different hemoglobinopathy which I will be discussing today specifically the variant hemoglobin before I go there let us have some quick review first. So tatandaan sa talasemia we will be going back to talasemia together with your extrinsic defects causing hemolysis in the finals. But now let's have a review. So this is a structure of hemoglobin structures of globin genes. So you all remember that we have six. We have six, your alpha we have your six. So the chromosome 16 codes for your alpha and your zeta you can found there your alpha or zeta genes on chromosome 16 and then you can also see your beta gamma delta epsilon on your chromosome 11 on your chromosome 11. And did you know that chromosome 16 chromosome 16 code for alpha and zeta alpha and zeta are collectively known as your alpha like genes and your chromosome 11 your beta gamma delta epsilon collectively these four are called your beta like genes. Your beta like genes. So I want you to remember alpha like genes and beta like genes because this will be important this are foundational when we go to your talasemia. Kasi kung ewan ko if you did have your advanced reading you would now know that there are two types of talasemia alpha and your beta talasemia and that is related to this too. So moving forward maybe some of you are wondering sir paano po nangyari yun no remember that in your remember that in your your fetal hemoglobins you have your goer 1 your goer 2 your portland and you have your hemoglobin F but some of you would be wondering sir paano po nangyari yun na from gamma from gamma from gamma naging beta na yung production from gamma which is in your goer 1 alpha and gamma in your goer 1 naging alpha and beta now becoming your hemoglobin A which is your adult hemoglobin so that is because of your BCL 11A and your crouple like factor 1 and your zinc finger transcriptional receptor necessary substances that silence your gamma globulin your gamma globin gene and are part of a complex mechanism involved in your gamma beta switching so they turn off your gamma gene they turn off their gamma gene and turns on turns on your beta gene that's why from producing goer 1 magiging production na tayo nang hemoglobin A hemoglobin A so that is just one thing that I want you to know so remember that this genes can actually undergo mutation they can have genetic mutation causing now different hemoglobinopathies different hemoglobinopathies and we have four ways on how genetic mutation would occur in your hemoglobinopathies would occur in your hemoglobinopathies we have your point mutation we have dilation insertion we also have your chain your fusion and in some cases your chain extension so let's go to the first one which is your point mutation your point mutation is actually the most common genetic mutation occurring in hemoglobinopathies so in point mutation there is a replacement of a nucleotide with a different nucleotide so if the AAA na na codon one of the nucleotide will be substituted for example magiging aga na lang magiging AGA na lang magiging AGA na lang magiging AGA na lang yung nucleot yung codon mo so what I'm trying to say here there is a point mutation only so meaning to say may mapapalitan na nucleotide so kuna re from take for example we have here your we do have here we do have the here your A take for example merang kang AGT merang kang aga so yung aga mo will now be mutated into becoming what they will now be mutated into your A sabi na natin AU A mag-mumotate siya to AUA take for example ganyan mapapalitan yung G na ng U so in that case we will now have your point mutation but the thing about point mutation is that continue pa rin yung reading frame and when I say reading frame the translation or the translation still continues okay so review your central dog maharis transcription and the translation in the reading frame it still continues so meaning to say may mapuproduce pa rin na may mapuproduce pa rin na globin or may mapuproduce pa rin na protein it's just that it will now be a defective a defective hemoglobin dahil sa iba na nga yung nucleotide na naipasok iba na nga yung nucleotide na naipasok therefore iba rin yung amino acid na naproduce after translation so that is for your point mutation indelation and insertion indelation there is a removal of one or more nucleotide in insertion there is a addition of one or more nucleotide in this case meron tayong disrupted na reading frame at kapag disrupted yung reading frame mo that will now cost your quantitative hemoglobinopathy or your tala simya okay na kukuha so madalas kapa ang ating hemoglobinopathies are actually point mutation and our delation to be specific causes your tala simya okay tala simya sir kasi instead ito kasi nagkukontinue so ibig sabihin nakakapagproduce ka pa rin ng globin abnormal man yung globin mo nakakapagproduce ka pa rin may hemoglobin pa din na napuproduce okay defective na lang indelation dahil mali or dahil kulang na yung nucleotide mo sa kodon anong nangyayare hindi na nakakapagproduce hindi na tayong nakakapagproduce ng hemoglobin therefore walading enough hemoglobin so nakakuha yung difference between point mutation and delation if you have other questions i will be entertaining them by the end of our discussion so having said that we also have your chain extension chain extension will just discuss it briefly so in chain extension this occur when the stop kodon is mutated so remember di ba meron tayong stop kodon your uga uwag your uwag so those are actually pala tandaan for your stop kodon we have your um we have your uwag we have your uwag and we also have your um uwag so those are your stop kodon uwag uwag those are your stop kodon so anong nangyari sa chain extension nag mutate yung stop kodon so parang si raya yung sabi na nating si raya yung stop light so hindi tumigil yung mga sasakyan ang nangyari nagtuloy-tuloy yung mga sasakyan all the way dahil wala ng stop light dun din sa translation wala ng stop kodon kaya tuloy-tuloy yung pagtranslate na amino acid chain we have a longer amino acid chain compared to the usual yun naman yung nangyari sa chain extension in your fusion there is a gene fusion gene fusion occur when two normal genes break between nucleotides switch position and then they anneal to the opposite gene so that is now pure fusion so these are just two of the genetic mutation but what I want you to remember guys are these two your point mutation and the deletion and insertion since we are talking about genetic mutation it's also rightful for us to talk about zygosity in zygosity this refers to the association between the number of gene mutation and the level of severity of the resultant genetic defect anong ibig sabihin natin dito sir zygosity refers to kung ilang gene the number of gene that is mutated because the number of gene mutated would directly affect would directly affect the severity of your symptoms the severity of your symptoms so always do remember always do remember that your globin genes are actually they came from from your mother and your father so one globin gene rather from your mother and one globin gene from your father okay so if both of those gene coming from your mom and your dad are both mutated if they are both mutated example natin yung beta gene both your beta genes are actually mutated then we call it homozygous homozygous we call that homozygous kapag naman isang gene lang yung abnormal or mutated yung samam mo lang or sa father heterozygous ang tawag natin don heterozygous if one gene is only mutated both the gene your mothers and your fathers gene are both mutated we call them homozygous okay anong relation ng heterozygosity and homozygosity pagdating sa severity ng symptoms at ng disease homozygous we call it the disease so this is the full blown disease in particular kapag heterozygous naman this will now be characterized by a mild sometimes asymptomatic na condition in some cases your patients with your take for example your hemoglobinopathy trait are eventually just carrier lalo na kapag asymptomatic naman sila carrier lang sila ng mutated gene so moving forward dahil nakabi ko na rin naman okay hopefully clear tayo dyan we have now different hemoglobin warians we have your hemoglobin SC C-HAR-L-M-E-O-R-AB D-PON-JAB-G in your G Philadelphia so all of this we will be discussing today so why do we have this yellow this different coding hemoglobin S is actually the most common and the most severe your hemoglobin C is the second most common hemoglobin E is the second most common hemoglobin variant okay your SC-E are the most common most common C-S second CC tapos C-E most severe severe S-S-S-severe so let's discuss now your hemoglobin variant before that let's distress inhale for one last time inhale exhale cold alright let's start our discussion about variant hemoglobin variant hemoglobin now we are actually transversing your hemoglobinopathies ito na yung iba-ibang hemoglobinopathies and remember there is a problem in the quality of your hemoglobin that is brought about by your point mutation okay by your point mutation so your hemoglobin hemoglobinopathies can either be a a disease or just a trait okay a disease or a trait depending on the of your gene so before we move forward into those specific hemoglobin variants let us talk about how can we differentiate one from the other right how can we differentiate one from the other so there are actually two major test one is your screening and the other one is your confirmatory test okay so they can be differentiated by their solubility test this is your screening test and in your solubility test we will be discussing two methods later which are your sodium disionite and your sodium betmetabysulfite test okay next is we also have your mobility in the electrophoretic field which is the diagnostic test okay in the diagnostic test we are using your electrophoresis okay we are using your electrophoresis and in this case we are using two halilose acetate which is 8.4 to 8.6 we also call it your alkaline or your alkaline electrophoresis and we also have your citrate electrophoresis which is your acidic electrophoresis that has a pH of 6.0 to 6.2 okay 6.0 to 6.2 so but first one let's talk about your sodium disionite test your sodium disionite test is a solubility test sir bakit po kailangan natin ng solubility test solubility test is to detect the solubility obviously of your hemoglobin originally your hemoglobin are soluble in your plasma but in cases of hemoglobinopathy some hemoglobins are actually insoluble in your some are actually insoluble so for your screening test we have your sodium disionite or your sodium hydro sulfite tube test okay this is a tube test so this is a screening test for the detection of your sickling hemoglobin so most especially kung mapapansin ninyo by this time we are talking about hemoglobin S or your sickle cell anemia so your hemoglobin S is actually the most severe that's why that is what we are trying to detect so your hemoglobin S or in your sickle cell anemia there's actually a formation of your sickle cell or sickle hemoglobin I mean sickle yung sickle napang lines ng mga damo yung hawak ng kamatayan that is a sickle shape yun yung sickle so in your RBC remember your RBC is a discoid by concave but in your sickle cell anemia in the presence of your hemoglobin S it becomes now sickle shape okay kaya usyutin awag na sickle cell anemia okay and what is present in sickle cell anemia is your hemoglobin S hemoglobin S so again in your sodium disionite this is a screening test detecting your sickling hemoglobin your hemoglobin S seen in your sickle cell anemia your sodium disionite is actually not specific so what we do here is first we lyse the cell to liberate the hemoglobin to liberate the hemoglobin so once the red blood cells are lyse using your saponin you add now your sodium disionite your sodium disionite binds and remove your oxygen from the test environment so once the oxygen is removed okay sickling will happen sickling will happen and what will happen next upon the removal of your oxygen your hemoglobin S will start to polymerize which is a result of the oxygenated state and will now form a precipitate in a high molarity phosphate buffer solution the precipitate consists of a tactoids which are RA liquid crystals okay which are RA liquid crystals this tactoids refracts and they deflect light that makes now the solution turbid so the positive result for your sodium disionite test is actually a positive turbidity kapag naging turbid yung solution after the addition of your sodium disionite positive positive result for your sickling anemia or your sickling cells sickling cells mamaya just hold on to your horses it will all come to realization later on okay next one we also have your sodium metabi sulfate okay your sodium metabi sulfate naman is actually a is done in your so what you do is you add your blood okay and you add 2% sodium metabi sulfate which is a reducing agent what happened here dahil nga may reducing agent it will now consume the oxygen there will be at the oxygenated state thereby your hemoglobin S will cost the formation of your sickling cell and then you will now look into it under your microscope and there will be now a formation of your sickling cell okay a formation of your sickling cell so sickling cells um or how well leaf how well leaf appearance will now cost a positive result kapag normal lang yung RBC or pag slightly granated sila negative sila so meaning to say there are no hemoglobin S um in the in the blood pero kapag nakaroon ka ng sickling cell formation in the form of your sickling cell or your how well leaf how well leaf formation okay how well leaf appearance this is now a positive result for your 2% sodium metabi sulfate test um ergo you have your hemoglobin S in them okay you have your hemoglobin S with them so I hope I'm making sense so we have to your sodium diethionite and your sodium metabi sulfate test so if we're gonna just summarize it both are screening tests um checking the presence of your hemoglobin S okay both are screening tests in the detecting or trying to screen if there is a presence of your hemoglobin S okay hemoglobin S so how do we confirm it so we have here your confirmatory test which are your electrophoresis so the mobility in the electrophoretic field will now differentiate your hemoglobin okay so we have here okay we have your hemoglobin molecules have a net negative charge okay in your alkaline pH that's why all of them will now try to migrate in your anode or your positively charged um electrode at cellulose acetate which is your alkaline electrophoresis at 8.4 to 8.6 the fastest is your hemoglobin H hemoglobin I the slowest is your C, E, O and your hemoglobin A2 kung mapapansin ninyo there are hemoglobin variants here but what I want to highlight is your the band where hemoglobin S, G and D are all together okay are all together generally this the other types of hemoglobin variants are generally mild okay most of them are generally mild the most severe or the highlight ng ating hemoglobin opatis for today will be your hemoglobin S your hemoglobin S which leads now to your sickle cell anemia and kung mapapansin ninyo sabi mo sir confirmatory test di ba to RUMOSAMIN that the confirmatory test should be specific but in this case hindi ko mahahanap si hemoglobin S bakit? kasi si hemoglobin S kasama niya si G and D ano itchura niyan kakawad nakita mo sa electrophoretic pattern magkakasama sila so hindi mo alam kung yung nagmigrate dito ay S ba? hemoglobin S ba? hemoglobin D ba? or hemoglobin G? so hindi ko po alam sir kung anong hemoglobin yung nandyan sabi nyo confirmatory test paano kung ngayon makokonfirm na may hemoglobin S doon ngayon papasok yung ating dito gamit natin si cellulose acetate correct? dito ngayon papasok yung ating citrate agar hemoglobin electrophoresis which is your acid electrophoresis and in this case citrate agar is used as a compliment remember it is used as a compliment to cellulose acetate electrophoresis sir, pag sinabing compliment gagawin mo lang to in complimentary hindi to replacement hindi to replacement so it differentiates your hemoglobin variant that migrate together and what are those hemoglobin variant these are your S, D, G and lepore all of these are hemoglobin variant but what you're trying to look for is your hemoglobin S kasi siya yung mong severe kasi siya yung magkokos ng sakit at siya yung magkokos ng complication siya yung magkokos ng sickle cell animya sa iyong paciente and that's what we're trying to look for and what are we going to use we're going to use citrate agar hemoglobin electrophoresis as a complimentary test in your as a complimentary test in your cellulose acetate electrophoresis so your citrate agar hemoglobin electrophoresis once again is used to differentiate your hemoglobin S from D and G hemoglobin S from D and G bakit po siya kailinawog na complimentary kasi kasi unang-unang-unang gagawin sa cellulose acetate you will now first do your cellulose acetate electrophoresis first and then kapag nakita mo na mayroong abnormal band kumakikita mo sir pa na mo naman na sabing abnormal because these are the normal patient this is the control control sa one patient 2, 3, 4, 6, 8 and 10 are all normal patient na mayroong band which you know are band for S, D, G, and lapore pero hindi mo nga alam kung S, D, G, or lapore ba yan kaya ang gagawin mo is your citrate agar hemoglobin electrophoresis and this is what will differentiate them in cellulose acetate mapapansin nyo S, D, G para parayat silang nandun sa iya isang band but here in your citrate agar remember the application point is in the middle application point is in the middle so you have the anode and you have the anode and you have the cathode your hemoglobin ang goal natin mahiwalay si S, D, G mahiwalay si S, D, G so take for example na apply ka dito ng ano makikita mo normal so pano po natin ma-dedeferensyate you will now be able to differentiate it by their mobility pattern ma-dedeferensyate mo si C at S dahil mas mabilay si C mas mabagal si S pero mas ma-dedeferensyate mo naman ang goal nga natin ma-dedeferensyate si S at si D and G kung mapapansin nyo hemoglobin D, hemoglobin G they migrated in the cathode hemoglobin S migrated in the anode so having said that now you will now start to confirm or you will now you will be able now to classify or identify hemoglobin S in your citrate agar electrophoresis because your hemoglobin S will migrate in the anode versus your hemoglobin D and your hemoglobin G that will migrate in the cathode naging gets back o yung hemoglobin S mo na differentiate mo kaya siya kaya D at G kasi C S pumunta sa anode pumunta sa cathode na hindi mo ma-identify kapag cellulose acetate kasi all of them will be migrating will be migrating in your anode all of them rather all of them will be migrating in your cathode dito sa cathode will migrate kasi negatively charged paray-paray at sila ng position dito nahiwalay mo na si S at nahiwalay mo na si D at si G again, alam ko nakakaano let's try to ponder upon all the things that have been said and done exhale hold inhale exhale hold and now let's move on to the different hemoglobin variants that we are going to talk about so the first one is your hemoglobin as ko mapapantinin nyo ino na nang sir Joms yung ating mga we actually discussed first the diagnostic test like your sodium abysulfite your sodium diethionite test and then your electrophoresis test for you to be able to differentiate the other hemoglobin variants the other hemoglobin variants why did I did that kasi we will now try to differentiate the different hemoglobin variants based on their solubility and their electrophoretic pattern so alam nyo na na si S, D, and G sama-sama sa A1 kaya kailangan mo ng complementarities which is your acid your citrate agar electrophoresis which will now differentiate your hemoglobin S from hemoglobin D and G so let's start with hemoglobin S so we all know that hemoglobin S causes your hemoglobin your sickle cell disease or your sickle cell anemia mamaya pagusapan natin yan so hemoglobin S has a structural defect so there is a mutation in your beta-globin in the beta gene there is a mutation in your beta-globin chain at possession 6 glutamic acid which was now replaced with your valine which was replaced by your valine ano pung nangyari ang glutamic acid mo napalitan ng valine so remember that in a normal beta chain gag for your glutamic acid and your sickle beta chain ang nangyari napalitan ng D in a sense nagkaroon tayo ng point mutation and that point mutation dahil sa instead na glutamic acid nandon sa possession 6 naging valine naging valine so hemoglobin solubility test they have a positive result setting positive result the one on your right with my laser and the one here is a negative a negative result for your solubility test pag sinabi kung solubility test this one dahil na sa test tube this is your sodium thionite test metabi sulfite pag nasa nasa glass slide so again remember this guys ha remember this guys so again your hemoglobin S both migrate in your cellulose acetate with your SDNG but we can separate that using your acid your acid citrate so again in hemoglobin S we have a mutation in betaglubin ganito natin siya babasayan there's a mutation in the betaglubin chain whereby at possession 6 your glutamic acid was replaced by your valine so this will now cause your sickle cell anemia ano ba yung sickle cell anemia your sickle cell anemia if it is a homozygous we will now call it a sickle cell anemia but if it is just a trait meaning to say minor lang yung symptoms or mild lang yung symptoms in your patient we have now your sickle cell trait your sickle cell trait and this under your sodium metabi sulfite test this is how your your sickle cell will look like so these are your hemoglobin these are your sickle cell these are red blood cells that contains your that contains your hemoglobin S sir bakit po sila naging ganyan again because of your because because of your valine because of your valine there is a structural changes kapag walang yung environment kapag walang oxygen na haawak yung mga cell na yan naging ganyan sila ok so kumakit kita niyo these are your sickle cells ok these are your sickle cell and the problem with your sickle cell ok bakit ba kasi galit na galit ka sa sickle cell sir ok bakit ba galit na galit ka sa sickle cell sir your sickle cell anemia so remember di ba the of your RBC gives it deformability deformability that enables it to pass through to pass through small capillaries anong nangyari kapag sickle cell kapag sickle cell they cannot deform ok they cannot they cannot deform themselves to pass through the vessel ang nangyari nag-accumulate sila yan nag-accumulate sila yan dahil nag-accumulate sila yan hindi sila nag-flow ok nag-babara yung blood vessels natin ok so this happens kapag walang oxygen so meaning to say sir kapag na oxygenate ulit sila babalik sila sa dati yes ok so kapag na oxygenate ulit sila babalik sila sa dati ok so by the way nice thing to know pakisulat na lang po sa inyong mga notes that your patient with sickle cell are actually immune to plasmodium falciparum ok they are they are actually immune to plasmodium falciparum wala kang plasmodium falciparum meron ka naman hemoglobin S so mamili ka na lang ok so of course ok your RBC in hemoglobin S are characterized to have sickle cell you can also see target cell every now and then since this is anemia you would eventually see nucleated RBC there is also cases of polychromasia bakit po may polychromasia sir dahil na po bakit po may polychromasia because it is a type of anemia so your body would try to produce or release reticulocytes earlier so please remember that in sickle cell anemia the hallmark feature of hemoglobin S is vaso occlusion or vaso occlusion pagbabara ng inyong mga kaugat-ugatan so this is specifically seen in your capillaries this is seen in your capillaries there are also episodes which collectively we call it crisis there is also anemia and patients with sickle cell anemia are actually much prone to bacterial infections such cost by SREUs your strep pneumoniae and your hemophilus influenzae your hemophilus influenzae so those are common bacterial bacterial bacterial infections seen in hemoglobin S so hemoglobin S doesn't just cause anemia but there are also other complications brought about by your sickle cell anemia again sickle cell anemia because of the presence of your hemoglobin S there is hemoglobin S because there is a mutation in your beta chain at position 6 whereby your glutamic acid is replaced by your valine so that is for your hemoglobin S I know we're nearing the 1R mark so I'll go straight with hemoglobin C the rest of the hemoglobin are now just easier to remember because we're just gonna talk about the point mutation that happened and their solubility characteristic so for your hemoglobin C there is a structural defect again in your beta beta-globin chain and this time it's still at position 6 whereby your glutamic acid this time is now replaced by your lysine kanina glutamic to valine that is sickle cell anemia hemoglobin C hemoglobin C on the other hand ang glutamic mo na palitan ng lysine so compared to hemoglobin S compared to hemoglobin S where the polymers are long and thin here hemoglobin C are short and thick crystals so meron kang hemoglobin C crystal so hemoglobin solubility test this is negative say it doesn't have turbidity so hemoglobin C disease kumakikita ninyo ang formation ng thick ang RBC that are short but are thick so they have hemoglobin C crystals inside them there is hemoglobin C crystals inside them again that is hemoglobin C naging lysine naging lysine so this is characterized generally by a mild splenomegaly and mild normocytic, normochromic anemia so mild lang CC so on the other hand we also have your hemoglobin C Harlem your hemoglobin C Harlem also known as hemoglobin C Georgetown okay so C Harlem and hemoglobin C Georgetown so you have a structural defect okay both structural defect natin is your six ang glutamic mo ang glutamic mo na palitan ulit ng valine double substitution ang meron sa C Harlem glutamic ang sub position six mo naging glutamic and in position at position 73 ang ang asparginine mo naging aspartic acid okay so again double substitution for hemoglobin C Harlem from glutamic to valine at position six also from asparginine to aspartic acid at position 73 okay so your hemoglobin C Harlem okay collectively hemoglobin here we call it your hemoglobin SC Harlem so this is a combination of your hemoglobin S and hemoglobin C okay hemoglobin S and hemoglobin C so makikita ninyo kanin na di ba si hemoglobin C and hemoglobin S natin so hemoglobin C is actually positive to your solubility test bakit po siya positive when I say positive meaning to say it's insoluble so meaning to say meron siya nagkaroon ka ng turbidity and in this case meron ka sa glutamic at siya ka sa glutamic at siya ka sa valine okay glutamic at siya ka sa valine kaya ka may positive result so the disorder is called hemoglobin SC Harlem because both your hemoglobin S and hemoglobin C are seen but sir is this more severe than hemoglobin S mas severe pa din si hemoglobin S mas severe pa din si hemoglobin S mas severe pa din si hemoglobin S let's go na to hemoglobin E hemoglobin E anong problem natin ke hemoglobin E hemoglobin E is a structural defect this time naman there is a substitution at position 26 okay kanina nung hemoglobin C parayasidan ng hemoglobin C glutamic to lysine pero ito since different position ibarin siya dahil iba yung position kahit parayas yung substitution iba yung disease kasi na hemoglobin C tignan nyo glutamic yung lysine pero sa position 6 at position E sa 26 naman siya mutation at position 26 nagkaroon ng point mutation glutamic naging lysine glutamic naging lysine so this are negative in your hemoglobin solubility test so you can see hemoglobin E disease or hemoglobin E trait depending on the if this homozygos hemoglobin E disease kapag trait lang hemoglobin E trait if it is just heterozygos and usually it's asymptomatic kapag hemoglobin E so patients with hemoglobin E disease are characterized with again mild anemia and mild splenomegety we also have here your hemoglobin O-arab hemoglobin O-arab is a beta-globin there's a mutation in your beta-globin specifically at position 121 whereby there's a replacement of your glutamic acid naging lysine siya 121 from glutamic naging lysine siya so moving forward this hemoglobin O-arab is also negative with your solubility test and this order associated here is your hemoglobin O-arab disease hemoglobin O-arab disease again characterized with mild splenomegety kumapapansin ninyo lahat mild lang except for hemoglobin S next we have here your hemoglobin D punjab your hemoglobin D punjab is the structural defect again a mutation in your beta-globin chain at position 121 where your glutamic acid was replaced by glutamin your glutamic acid was replaced by your glutamin solubility test this are negative the disorder associated here is your hemoglobin D disease last but not the least we also have here your hemoglobin G and G Philadelphia here the problem is now not on the beta but on the alpha there is a structural defect on your alpha-globin chain specifically on position 68 whereby your asparginin was replaced with your lysine asparginin was replaced with your lysine at position 68 so there is a negative solubility test for this one and the disorder the disorder associated here is your hemoglobin G disease again characterized with a mild hemolytic anemia and mild splenomegaly so kung mapapansinin nyo once again everything are just mild except for hemoglobin S other hemoglobin variants we have the following your hemoglobin we have other hemoglobin variants so do we have to memorize this just the one that are discussed specifically so all of this are examples of substitution we have deletion your gun hill that is an example of deletion your hemoglobin constant spring is a chain extension kung mapapansin ninyo chain extension so again to sum it up we have different hemoglobins that we did talk about hemoglobin S, C, Harlem, E, O, R, D, P, N, G all of this you need to remember specifically the point mutation that had happened anong position, anong amino acid yung na ipalep so eventually we are going to differentiate them kung mapapansin ninyo hemoglobin S hemoglobin C, S, C, Harlem lang ang positive sa solubility test the rest are negative the rest are negative and of course remember their electrophoretic pattern and not only that remember your cideroblastic anemia your porphyria and how I differentiated your hemoglobinopathies and your talasemia with that thank you so much for listening and if you have any questions we will now proceed with the discussion with the question and answer portion again thank you so much for listening this has been Sir Jones and thank you for listening so I'll see you guys on our live question and answer portion see you there